• Cells are the fundamental units of all living organisms
• Each cell is a complex system consisting of many substructures
• Types of organisms:
  • Viruses are simplest organisms (∼ 10,000 bp. long genomes), which require a living host
  • Prokaryotes are simplest free-living organisms, e.g. bacteria (∼ 1,000,000 bp. long genomes)
  • Eukaryotes have cells which contain internal structures such as a nucleus, e.g. yeast
  • Multi-celled organisms involve cell specialization, requiring differential gene expression and inter-cellular signaling

• Chromosomes are packets of compressed and entwined DNA and are located in the nucleus.

• Each chromosome carries its own unique set of genes. The specific site along a specific chromosome that a gene is located is called its genetic locus.

• Humans have a total of 46 chromosomes: 44 autosomes and 2 sex chromosomes. Autosomes occur in pairs - diploid.

• Because there are two copies of each autosome, there are 2 alleles of a gene at each locus.

• Germ cells, sperm and egg, carry only a single copy of each chromosome and gene and are called haploid. ## Centromere locations

• Centromere divides chromosomes into a short arm (p; petit) and a long arm (q)

• Centromere is defined by the first band on the short arm (p10) and the first band on the long arm (q10)

• Telomeres are at the termini (pter and qter)

• Cytogenetics is the study of normal and abnormal chromosomes.
• The normal configuration of chromosomes is often termed the euploid karyotypic state.
• Euploidy implies that each of the autosomes is present in normally structured pairs and that the X and Y chromosome are present in normally structure pairs for the sex of the individual.
• Deviation from the euploid karyotype - the state termed aneuploidy - is some alteration in the overall chromosome structure, such as loss of entire chromosomes, the presence of extra copies of chromosomes, etc.

• Genes are discrete hereditary units located on the chromosomes (DNA).
• Each gene provides a clear and unambiguous set of instructions for producing some property of its organism.
• The complete set of genes in an organism is referred to as its genome.

• The basic unit (nucleotide) is composed of an organic base attached to a deoxyribose sugar
• The phosphate group also attached to the sugar
• The base is one of cytosine (C), thymine (T), adenine (A), and guanine (G)

• In the mid-1900’s methods were developed to size separate and visualize DNA within an electrically charged gel (originally made from sugar)
  
• DNA is loaded into different “lanes” at the top of the gel, and the charge is applied
  
• Key idea is that smaller fragments of DNA would move faster through the gel and be towards the bottom than longer fragments that will be towards the top.
  
• Allows to test for presence/absence of DNA as well as compare relative lengths of molecules
  
• Will often reserve one of the lanes for a “DNA ladder” with fragments of a known size distribution

• DNA is a double helix, with bases to the center (like rungs on a ladder) and sugar-phosphate units along the sides of the helix (like the sides of a twisted ladder).
• The strands are complementary (Watson-Crick base pairing rules)
• A (purine) pairs with T (pyrimidines) C (pyrimidines) pairs with G (purine)
• The pairs held together by hydrogen bonds. The helix is caused by the use of the hydrogen bonds between the single-strands

• The force that holds a base pair together is a weak hydrogen bond.
• Although each individual bond is weak, their cumulative effect along the strands is strong enough to bind the two strands tightly together.
• As a result, DNA is chemically inert and is a stable carrier of genetic information.

• In DNA replication, the DNA molecule unwinds and the “ladder” unzips, thereby disrupting the weak bonds between the base pairs and allowing the strands to separate.
• Nucleotides have to be assembled and available in the nucleus, along with energy to make bonds between nucleotides.
• DNA polymerases unzip the helix by breaking the H-bonds btw bases
• Once the polymerases have opened the molecule, an area known as the replication bubble forms (always initiated at a certain set of nucleotides, the origin of replication).
• New nucleotides are placed in the fork and link to the corresponding parental nucleotide already there (A with T, C with G).

• Amazing biotechnologies for manipulating DNA molecules are used as building blocks for even more powerful technologies.
• DNA synthesis machines enable one to grow short DNA molecules of a specified sequence.
• The Polymerase chain reaction (PCR) enables one to make many copies of a particular DNA sequence anywhere in solution given only the starting/ending sequences (primers).

• Developed in 1983 by Kary Mullis, PCR allows for the amplification of DNA fragments that are flanked by known “primer” sequences.
• Exquisitely sensitive and specific, can amplify a single molecule in a sample into billions of copies with nearly perfect fidelity
• Uses naturally occurring polymerase enzymes that copy DNA by adding free nucleotides to a single-stranded template

• Messenger RNA (mRNA) - carrier of genetic information
• Transfer RNA (tRNA) - deliver amino acids for protein synthesis
• Ribosomal RNA (rRNA) - central component of ribosome, protein manufacturing machinery
• Small RNA (siRNA, miRNA, snRNA, piwiRNA) - regulation of transcription/translation

• In transcription, the DNA double helix opens along its length
• One strand of the open helix remains inactive, while the other strand acts as a template against which a complementary strand of mRNA forms
• The sequence of bases along the mRNA strand is identical to the sequence of bases along the inactive DNA strand, except uracil (U) replaces T. Also RNA has ribose sugar instead of deoxyribose sugar.
• RNA (single-stranded) moves out into the cytoplasm.

• RNA polymerase is an enzyme that produces RNA

1. RNA Pol I - transcription of ribosomal RNA (not the 5S subunit)
2. RNA Pol II - mRNA, snRNA, microRNA
3. RNA Pol III - tRNA, 5S rRNA, small RNA

• Non-coding interruptions are known as intervening sequences or introns.
• Coding sequences that are expressed are exons.
• Most, but not all structural eukaryote genes contain introns. Although transcribed, these introns are excised (cut out) before translation.

• Exon - EXpressed regiON
• Intron - INTragenic regiON
• mRNA splicing - variants of mRNA assembly

• Tissue specific alternative splicing patterns of the \(\alpha\)-tropomyosin pre-mRNA molecule
  
• Exons are blue rectangles
  
• Introns are black carets

• After eukaryotes transcribe an RNA, the RNA transcript is extensively modified before export to the cytoplasm.
• A cap of 7-methylguanine (a series of an unusual base) is added to the 5’ end of the mRNA. This cap is essential for binding the mRNA to the ribosome.
• A string of adenines (as many as 200 nucleotides known as poly-A) is added to the 3’ end of the mRNA after transcription. The function of a poly-A tail is not known, but it can be used to capture mRNAs for study.
• Introns are cut out of the message and the exons are spliced together before the mRNA leaves the nucleus.

• Each cell contains a complete copy of the organism’s genome. A gene that is transcribed is said to be expressed
• Not all cells express the same genes which is why different cells perform different functions
• Even within the same cell different genes will be expressed at different times and perhaps at different levels

• Housekeeping genes are genes that are required for the maintenance of basal cellular functions that are essential for the existence of a cell, regardless of its specific role in the tissue or organism.
• They are expected to be expressed in all cells of an organism under normal conditions, irrespective of tissue type, developmental stage, cell cycle state, or external signal.
• Can be used as internal controls in gene expression studies

• Typical examples:
  • glyceraldehyde- 3-phosphate dehydrogenase (GAPDH)
  • tubulins (beta-tubulin TUBB)
  • cyclophilin (cyclophilin A CYPA)
  • albumin (ALB)
  • actins (beta-actin ACTB)
  • 18S rRNA or 28S rRNA.

• Should they be expressed at constant level?
• How to account for alternative splicing?

3,804 housekeeping genes (and exons) defined from Human BodyMap project gene expression data http://www.tau.ac.il/~elieis/HKG/

• Computational biology attempts to use genome sequence to ascertain function of genes.
• Although genomes vary slightly from person to person, it seemed reasonable to try to establish a consensus human genome sequence.
• Robert Sinsheimer, chancellor of UC Santa Cruz, proposed to sequence the human genome in 1984.
• After much debate, the human genome project started in October 1990.

• Mendel’s work implied that the entire repertoire of an organism’s genetic information - its genome - is organized as a collection of discrete, separable information packets, now called genes.
• His research implied that the genetic constitution of an organism (its genotype) could be divided into hundreds, perhaps thousands of discrete information packets
• The observable outward appearance of an organism (its phenotype) could be subdivided into a large number of discrete physical or chemical traits.

• genotype: The genetic (alleleic) makeup of an organism with regard to an observed trait. The sum total of sequence variations (polymorphisms and mutations) present in a genome.
• phenotype: The observed properties or outward appearance of a trait.

• Mendel’s Second Law (The Law of Independent Assortment) states alleles of one gene sort into gametes independently of the alleles of another gene.
• However, Morgan and his student Sturtevant noticed that for certain traits the probability of having one trait given another was not 50/50 – those traits are genetically linked

• Sturtevant realized the probabilities of co-occurrences could be explained if those alleles were arranged on a linear fashion: traits that are most commonly observed together must be locates closest together
• Today genetic maps are routinely generated by measuring the rates of polymorphic markers in large populations of individuals

• Though DNA is stable, the genome is corruptible, in other words, the genetic code can be changed.
• An allele that is present in the great majority of individuals within a species is termed wild type (naturally present in large numbers of apparently health organisms).
• Mutations are when one allele is converted into another allele or an allele is created. The collection of alleles present in the genomes of all members of a species is the gene pool for the species.

• The two copies of a gene could convey different, possibly conflicting information. The different versions of a gene is called an allele.
• Organisms with two identical alleles of a gene are homozygous
• Organisms with two different alleles of a gene are heterozygous.

• When a gene is heterozygous, the observed phenotype encoded by one allele of a gene is dominant with respect to the phenotype encoded by another allele, the recessive one.
• The alleles of some genes may be co-dominant, wherein a blend of the two alleles result in a phenotype.
• Incomplete penetrance is when a dominant allele is present but the phenotype is not manifested because of the actions of other genes in the organism’s genome.

• Autosomal dominant
• Autosomal recessive
• X-linked dominant
• X-linked recessive
• Mitochondrial
• Non-Medelian (e.g., imprinting)

• Evolutionary change happens because of changes in genomes due to mutations and recombination.
• Mutations are rare events, sometimes single base changes, sometimes larger events.
• Recombination is how your genome was constructed as a mixture of your two parents.
• Through natural selection, favorable changes tend to accumulate in the genome.

• Evolution motivates homology (similarity) search, because different species are assumed to have common ancestors. Thus DNA/amino acid sequences for a given protein (e.g. hemoglobin) in two species or individuals should be more similar the closer the ancestry between them.
• The genetic variation between different people is surprisingly small, perhaps only 1 in 1000 base-pairs.
• Homology searches can often detect similarities between extremely distant organisms (e.g. humans and yeast).

• Phylogenic trees based on gene homologies provide an independent confirmation of those proposed by taxonomists. This is convincing evidence of evolution.
• A host of interesting computational problems arise in trying to reconstruct evolutionary history.

• One base being replaced by another (substitution)

• A base being excised (deletion)

• A base being added (insertion)

• A small subsequence of bases being removed and reinserted in the opposite direction (inversion)

• A small subsequence of bases being removed and reinserted in a different place (translocation)

• Since DNA is information, and information typically has a beginning point, an inversion would produce an inactive or altered protein.

• Likewise deletion or duplication will alter the gene product.

• Transmission of a mutation from one generation to the next, by the germ cells (sperm and egg), is said to occur via the germ line.
• Mutations affecting the genomes of cells everywhere else in the body, which constitute the soma, have no prospect of being transmitted to offspring and are called somatic mutations.

Chromosome disorders
- Congenital (7 per 1000 newborns, 50% of spontaneous first trimester abortions)
- Acquired (cancer)

Single-gene disorders Individually rare
- As a group affect \(\sim\) 2% of population over lifespan

Multifactorial or complex disorders
- A result of combination of genes
- May affect \(\sim\) 60% of entire population